Date of Award

9-2010

Document type

Thesis

Degree Name

MSc by research (Master of Science by research)

First Supervisor

Dr Jacqueline S. Daly

Keywords

Bone Diseases, Hip Fractures, MicroRNAs

Abstract

Bone is a dynamic material critical to human form and function. Bone strength and structure is maintained by a dynamic homeostasis; bone re-modelling. Osteoporosis is a degenerative bone disease whereby reduced bone quantity and quality results in increased fracture risk. Osteoporotic fractures confer high levels of morbidity and mortality. Osteoporosis is escalating in its impact worldwide in parallel with the growing global population and increasing life expectancy affecting both individuals and the health care systems that serve them. Novel diagnostics, prognostics and therapeutics are needed for osteoporosis. miRNA are small non-coding RNAs which achieve a regulatory function by complimentary binding to the 3' UTR of target mRNA to bring about their translational inhibition or degradation, thus regulating gene expression. miRNA have been shown to have a role in many body systems and diseases and have previously been associated with bone dynamics and disease in-vitro.

A method was designed and optimised for the processing, homogenisation and reliable extraction of good quality total RNA from fresh frozen bone tissue. Amplification of rniRNA from this total RNA was achieved. The miRNA profiles of trabecular and cortical femoral bone tissue from osteopenic and osteoporotic hip fracture patients were assessed and compare to controls. miRNA consistently differentially expressed with disease were short-listed and cross referenced to identify their predicted mRNA targets of interest in bone dynamics and disease. 76 miRNA were demonstrated to be consistently differentially expressed in diseased bone tissue, with most being specifically differentially expressed in either cortical or trabecular bone. Only 5 miRNA were differentially regulated in both bone types. The differential expression of miRNA with disease state indicates a role for miRNA in bone dynamics and disease, with miRNA short-listed linked to predicted target mRNA previously shown to be key in osteoblast and osteoclast differentiation and proliferation.

Two of the miRNA short-listed by profiling, miR-532-3p and miR-877, were specifically experimentally up- and down-regulated in a human osteoblast cell line. The promotion and suppression of these two miRNA was shown to have no effect on alkaline phosphatase and collagen type 1 A1 expression levels over the course of a 6 day study in-vitro.

Creative Commons License

Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-Share Alike 3.0 License.

File Size

11,890 KB

Comments

A thesis submitted to the Royal College of Surgeons for the degree of Master of Science, 2010.

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